The mechanism by which protein toxins kill cells is quite complex. Many toxins bind to receptors on the surface of mammalian cells, are internalized by endocytosis, translocate to the cytosol and there exert an enzymatic activity that kills the target cell. Accordingly, these toxins have separate domains for cell binding, translocation and an enzymatic activity that lnactivates an essential cellular function. Pseudomonas exotoxin-A (PE) is a single polypeptide chain of 613 amino acids. X-ray crystallographic studies and mutational analysis of the PE molecule have shown that PE consists of three domains: an amino terminal cell receptor binding domain (Domain I); a middle translocation domain (Domain II); and a carboxyl terminal activity domain (Domain III). Domain III catalyzes the ADP ribosylation and inactivation of elongation Factor 2(EF-2) which inhibits protein synthesis and leads to cell death. Mutational analysis of Domain I has revealed that lysine.sup.57 plays a major role in receptor binding. Similarly glutamic acid.sup.553, Tyrosine.sup.481 and histidine.sup.426 have been shown to be important for ADP-ribosylation activity. Recently mutational analysis of domain II has shown that certain portions of this domain are absolutely required for the cytotoxicity of PE.
While constructing various chimetic toxins in which growth factors were fused to a form of PE (PE40) which was devoid of domain I, it was observed that the recombinant fusion proteins, made by attaching TGF.alpha., interleukin-2 or interleukin-4 at the carboxyl end of PE40 had poor cytotoxic activity. Hence, an examination of the role of the carboxyl terminus of the PE molecule (domain III) was undertaken.